Vesicles are single bilayer shells which are composed of amphipathic molecules such as surfactants or detergents. As used in this disclosure, the term vesicle will be understood as referring to unilamellar bilayer shells having a relatively small size, for example, having a diameter less than about 200 nanometers. The term "liposome" should be distinguished for purposes of this disclosure as referring to multilamellar shells which are generally larger in size, e.g. up to several microns in diameter.
Vesicles have a number of important utilities, including chemical and biochemical applications. For example, vesicles are useful in performing biochemical assays which involve the storage or encapsulation of biological materials such as enzymes or their substrates, which allows for controlled protection and release of an encapsulated substance. It is also possible to incorporate a reaction substrate into a vesicle membrane bilayer for presentation on the surface of the vesicle. Both vesicles and liposomes are of considerable interest in the controlled release and targeted delivery of pharmaceutically active agents. Loading of, for example, a medication, into vesicles or liposomes can serve to protect the load from degradation or dilution in the blood. Vesicles are also useful in preparing models for the study of photosynthesis and membrane phenomena, by incorporating the appropriate molecules into the vesicle membrane in order to induce electron transfers and/or establish proton gradients.
Vesicles and liposomes have conventionally been prepared using complex amphipathic molecules and/or elaborate preparation techniques. For example, liposomes may be composed of structurally complex phospholipids of the type which are found in natural membranes, such as phosphatidyl choline or phosphotidyl ethanolamine. Complex phospholipids normally contain more than one non-polar hydrocarbon "tail" region on each molecule. Liposomes formed therefrom can also incorporate other naturally-occurring membrane substances, for example steroids such as cholesterol or their analogs and derivatives, or can be modified by the presence of a surfactant or polymer.
Non-phospholipid vesicles have been prepared from various surfactants and detergents, but their preparation in aqueous solution has required considerable mechanical energy and/or elaborate chemical treatments. For example, it is known that vesicles may be prepared in aqueous solution using sonication or pressure filtration. Chemical treatments for inducing vesicle formation include detergent dialysis and reverse-phase evaporation. Vesicle solutions prepared by sonication are metastable, however, and the vesicles revert over time to more thermodynamically stable structures, such as multilamellar liquid-crystalline aggregates. See, generally, Madani and Kaler, Langmuir 6:125 (1990). In the course of vesicular breakdown, the contents of the vesicles are released.
A currently popular class of surfactant materials which have been used to prepare surfactant vesicles include polyoxyethylene esters, ethers, and amines. Exemplary disclosure is found in U.S. Pat. Nos. 4,217,334; 4,670,185; 4,743,449; 4,853,228; and 4,911,928.
It is known that many surfactants, and most simple detergents, are capable of forming micellar structures and emulsions in aqueous solution. Many surfactants are commercially important emulsifiers. Single-tailed amphiphiles, having relatively small tail groups and large head groups, invariably form micelles in solution.
In the past, addition of a cationic surfactant to an anionic micellar solution has been largely avoided. Solutions of anionic and cationic surfactants may form a lamellar phase, or may form precipitates, when combined in equimolar amounts. Although from theoretical considerations it could be predicted that amphiphilic molecules with relatively large hydrophobic groups and relatively small hydrophilic groups are capable of forming bilayer structures, spontaneous vesicle formation in vitro is in fact only rarely observed. Moreover, mixtures of simple anionic and cationic detergents are expected to produce mixed micelles in solution. See, Chen and Hall, Colloid. Polym. Sci. 251:41 (1973); Barker et al., J. Chem. Soc. Faraday Trans. 1 70:154 (1974); J. F. Scamehorn, Ed., Phenomena in Mixed Surfactant Systems (American Chemical Society, Washington, D.C. 1986).
There have been reports of spontaneous vesicle formation in certain mixtures of short- and long-chain, double-tailed lecithins. See Gabriel and Roberts, Biochemistry, 23:4011 (1984). Spontaneous vesicle formation has been reported to take place in solutions of double-tailed surfactants with hydroxide and other more exotic counterions. See Talmon et al., Science 221:1047 (1983); Brady et al., J. Am. Chem. Soc. 106:4279 (1984). Vesicle formation has been reported in some heated mixtures of anionic and cationic monoalkyl surfactants, but the vesicles were reported to be only transiently stable, and degenerated rapidly on cooling. See, Hargreaves and Deamer, Biochemistry 17:3759 (1978). Although these systems reflect improvements over conventional sonicated vesicles, the relatively restricted chemical or physical properties of the vesicles, or the limited availability of the constituent surfactants, were such that these methods have not been widely exploited.